Mass spectrometry (MS) has long been a widely accepted analytical technique for obtaining qualitative and quantitative information from a sample. MS is commonly used to determine molecular weight, identify chemical structures, and accurately determine the composition of mixtures. MS is becoming increasingly important in biological research to determine the structure of organic molecules based on the fragmentation pattern of ions formed when sample molecules are ionized.
A well-known analytical technique which combines a separation technique with an analytical detection device is gas chromatography-mass spectrometry (GC-MS). In this method, GC can provide separations of sufficiently volatile compounds which are then ionized and analyzed by mass spectrometry. GC-MS has become established as the definitive analytical technique for amenable compounds, i.e., compounds having sufficient volatility for GC separation and ionization by conventional gas phase electron impact or chemical ionization methods used in mass spectrometry. Such an established capability of broad application is not known to exist for nonvolatile compounds and mixtures.
The coupling of mass spectrometers with liquid chromatography systems has provided a very valuable tool for identifying organic compounds. The unique value of the liquid chromatographic separation systems is their ability to separate solutions containing mixtures of organic compounds into liquid fractions containing individual compounds. However, the product of the liquid chromatographic column is an eluant liquid solution of the compound or compounds to be analyzed that is at atmospheric pressure, whereas the mass spectrometer analyzes compounds in a high vacuum system. Evaporation of the eluant solvent and presentation of the desolvated particles to the mass spectrometer in a suitable form has presented serious difficulties limiting the sensitivity of the mass spectrometer and greatly complicating its efficient operation. Currently Particle Beam (PB) and Electrospray Atmospheric Pressure Ionization (ES-API) Liquid Chromatography/Mass Spectrometers (LC/MS) are the two most popular interface techniques.
In ES-API LC/MS, an electrospray nebulizer produces an aerosol of charged droplets at atmospheric pressure from which desorbed charged analytes are separated. These ions are then electrostatically driven through a multistage separator in which the final pressure is reduced to about 10.sup.-5 torr.
ES-API is a soft ionization technique. For low molecular weight compounds it typically produces singly charged molecular ions and simple spectra. One feature of ES-API is its ability to produce multiple charged ions for high molecular weight compounds; this makes ES-API suitable for analysis of compounds with molecular weights far in excess of the nominal mass range of a quadrupole analyzer. In most designs, by adjusting specific potentials, structurally significant fragmentation can be produced via CID (collision induced dissociation) at the interface. ES-API is best suited for polar compounds, particularly high molecular weight compounds. Current interfaces operate at relatively low flow rates (&lt;100 .mu.l/min), although concentration dependency implies that higher flow rates could be split without loss in sensitivity. Most ES-API designs use a multistage pressure reduction system; some designs employ a large capacity cryopumping mechanism.
In PB LC/MS systems, an aerosol consisting of helium dispersion gas and droplets containing relatively low levels of analyte dissolved in LC effluent is first generated by a nebulizer. The aerosol is injected into a heated desolvation chamber where the volatile components of the droplets (primarily HPLC effluent) are vaporized, resulting in a mixture of helium gas, solvent vapor, and desolvated analyte particles. This mixture then enters a two stage momentum separator in which the less massive components (such as solvent vapor and helium gas) are pumped away while the more massive particles continue through the system and into the mass spectrometer source where the particles are vaporized, ionized, and mass analyzed. The momentum separator also serves as a pressure reduction and sample enrichment device, since most of the gas and solvent are pumped away, while most of the sample enters the mass spectrometer.
Particle Beam LC/MS is capable of producing classical, library searchable electron impact (EI) or chemical ionization (CI) spectra. Since PB relies on gas phase ionization methods, samples must have some degree of volatility. Although this may be significantly less volatility than required for GC/MS analysis, totally non-volatile samples cannot be analyzed. Furthermore, even those samples that have sufficient volatility may be too thermally labile for EI analysis. In this case, the use of CI can extend the analytical usefulness at the cost of structural information. PB LC/MS is best suited to non-polar compounds with molecular weight less than 1000 amu and low volatility.